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Evaluation of Antitumor Activity of Cuscuta Reflexa Roxb (Cuscutaceae) Against Ehrlich Ascites Carcinoma in Swiss Albino Mice

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dx.doi.org/10.4314/tjpr.v10i4.10 Abstract Purpose: The aim of this study was to investigate the antitumor effect of the chloroform and ethanol extract of the whole plant of Cuscuta reflexa Roxb. (Cuscutaceae) in Swiss albino mice against Ehrlich Ascites Carcinoma (EAC) cell line. Methods: The antitumor activity of the chloroform and ethanol extracts of Cuscuta reflexa was evaluated against Ehrlich ascites carcinoma (EAC) tumor in mice at doses of 200 and 400 mg/kg body weight orally, respectively, while acute oral toxicity studies were performed to determine the safety of the extracts. Briefly, the EAC cells were injected (i.p.) into ninty six mice (divided into 6 numerically equal groups), and after a one-day incubation period, the extracts were administered to the mice daily for 16 days. On day 21, six animals in each group were sacrificed for observation of antitumor activity and the remaining animals were observed to determine host the life span. Antitumor effect was determined by evaluating tumor volume, viable and nonviable tumor cell count and hematological parameters of the host. The standard antitumor used was 5-fluorouracil. Results: Administration of the extracts resulted in a significant (p < 0.05) decrease in tumor volume and viable cell count, but increased non-viable cell count and mean survival time, thereby increasing the life span of the tumor-bearing mice. Restoration of hematological parameters -red blood cells (RBC), hemoglobin, white blood cells (WBC) and lymphocyte count -to normal levels in extract-treated mice was also observed. Conclusion: The results suggest that the chloroform and ethanol extracts of C. reflexa exhibit significant antitumor activity in EAC-bearing mice that is comparable to that of the reference standard, 5-fluorouracil.
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Chatterjee et al
Trop J Pharm Res, August 2011;10 (4):
447
Tropical Journal of Pharmaceutical Research August 2011; 10 (4): 447-454
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http://dx.doi.org/10.4314/tjpr.v10i4.10
Research Article
Evaluation of Antitumor Activity of Cuscuta Reflexa
Roxb (Cuscutaceae) Against Ehrlich Ascites
Carcinoma in Swiss Albino Mice
Dandopani Chatterjee
1*
, Ram K Sahu
1
, Arvind K Jha
2
and
Jaya Dwivedi
3
1
Department of Pharmacognosy, Oriental College of Pharmacy, Bhopal (M.P.)-462021,
2
Shri Shankaracharya
Institute of Pharmaceutical Science, Junwani, P.O. Nehru Nagar, Bhilai, (C.G.)-490020,
3
Department of
Pharmaceutical Chemistry, Banasthali University, P.O. Banasthali Vidyapith (Rajasthan)-304022, India
Abstract
Purpose: The aim of this study was to investigate the antitumor effect of the chloroform and ethanol
extract of the whole plant of Cuscuta reflexa Roxb. (Cuscutaceae) in Swiss albino mice against Ehrlich
Ascites Carcinoma (EAC) cell line.
Methods: The antitumor activity of the chloroform and ethanol extracts of Cuscuta reflexa was
evaluated against Ehrlich ascites carcinoma (EAC) tumor in mice at doses of 200 and 400 mg/kg body
weight orally, respectively, while acute oral toxicity studies were performed to determine the safety of
the extracts. Briefly, the EAC cells were injected (i.p.) into ninty six mice (divided into 6 numerically
equal groups), and after a one-day incubation period, the extracts were administered to the mice daily
for 16 days. On day 21, six animals in each group were sacrificed for observation of antitumor activity
and the remaining animals were observed to determine host the life span. Antitumor effect was
determined by evaluating tumor volume, viable and nonviable tumor cell count and hematological
parameters of the host. The standard antitumor used was 5-fluorouracil.
Results: Administration of the extracts resulted in a significant (p < 0.05) decrease in tumor volume and
viable cell count, but increased non-viable cell count and mean survival time, thereby increasing the life
span of the tumor-bearing mice. Restoration of hematological parameters - red blood cells (RBC),
hemoglobin, white blood cells (WBC) and lymphocyte count - to normal levels in extract-treated mice
was also observed.
Conclusion: The results suggest that the chloroform and ethanol extracts of C. reflexa exhibit
significant antitumor activity in EAC-bearing mice that is comparable to that of the reference standard, 5-
fluorouracil.
Keywords: Cuscuta reflexa, Ehrlich ascites carcinoma, 5-Fluorouracil, Tumor volume, Viable cell count.
Received: 10 October 2010 Revised accepted: 22 May 2011
*Corresponding author: E-mail: chatterjee_dp@yahoo.com; Tel: +91-9893093497
Chatterjee et al
Trop J Pharm Res, August 2011;10 (4):
448
INTRODUCTION
Malignancy is one of the most serious
diseases afflicting mankind today and indeed,
the second most deadly disease after heart
disease. According to WHO estimates, more
than 7 million people died of cancer in
2005[1]. There exists a close relationship
between the occurrence, growth and decline
of tumor. In recent times, the demand for
more effective and safer therapeutic agents
for the chemoprevention of human cancer
has increased. Natural products produced by
plants and their synthetic derivatives are
expected to play an important role in the
development of innovative agents to inhibit
the onset of cancer [1]. There is a growing
interest in the pharmacological evaluation of
various plants used in the Indian traditional
system of medicine.
Cuscuta reflexa (Cuscutaceae), known in
Hindi as Amarbela, is a phanerogamic stem
parasite, and is distributed worldwide. The
species are rootless, leafless twining annual
parasites with yellowish stems, distributed
across tropical and temperate regions, and in
India about 6 species are found. It grows on
thorny or other shrubs, sometimes completely
covering bushes and trees [2]. C. reflexa
spread from one host to another, and on
each victim, they twine and cling tightly with
special branching organs called houstoria,
penetrating the host and connecting to the
host xylem as well as to the host phloem, and
absorbing from it both water and elaborated
food stuffs such as sugars and amino acids
[3]. Various parts of this plant are used in
tribal medicine for the diseases such as
impotence, premature ejaculation, sperm
leakage, frequent urination, ringing in the
ears, lower back pain, sore knees,
leucorrhea, dry eyes, blurred vision, and tired
eyes.
Cuscuta is one of nine herbs included in the
manufacture of Equiguard, a Chinese herbal
medicine recommended for kidney and
prostate disorders. Research performed at
New York Medical College indicates that the
combination of ingredients in Equiguard may
well be effective in the treatment of prostate
cancer. The preparation inhibited the growth
of cancer cells, increased the rate of self-
destruction (apoptosis) of cancer cells, and
prevented the surviving cells from forming
colonies [4]. Phytochemical investigation of
Cuscuta reflexa indicates the presence of
kaempferol-3-O-glucoside, astragallin [5],
myrecetin, benzopyrones [6], glucopyra-
nosides [7], propenamide, flavonols [8],
quercetin and quercetin-3-O-glucoside, β-
sitosterol, and bergenin [9].
The objective of the present study was to
evaluate the antitumor effect of Cuscuta
reflexa against Ehrlich’s Ascites Carcinoma
(EAC) in Swiss albino mice.
EXPERIMENTAL
Plant materials
Whole plants were collected from PNT
Square, Jawahar Chowk, Bhopal (M.P.)
India, during the months of January and
February 2007. The species was identified by
the local people during the time of collection
and later authenticated by Dr Pradeep Tiwari,
taxonomist, Department of Botany, Dr. H.S.
Gour University, Sagar (M.P.), India. A
voucher specimen (no. BOT/412/123) was
prepared and preserved along with the crude
drug sample at the herbarium of Department
of Botany, Dr. H.S. Gour University, Sagar
(M.P.), India. The whole plant was shade-
dried, reduced to coarse powder and stored
in an airtight container until further use.
Preparation of extract
The powdered drug (1 kg) was packed in a
Soxhlet apparatus and extracted for 24 h
with petroleum ether (60 - 80 ºC) to defat the
drug. The defatted material was then
extracted with chloroform (2.5 L). The
chloroform extract was separated while the
marc was extracted with ethanol (2.5 L, 95
%). The solvent, in each case, was removed
by evaporation and the residual solvent
Chatterjee et al
Trop J Pharm Res, August 2011;10 (4):
449
eliminated under reduced pressure. Separate
solutions of the chloroform and ethanol
extracts were prepared in two doses - 200
and 400 mg/kg - for the antitumor test.
Experimental animals
Swiss albino mice, weighing (23 – 25 g),
were used in this study. They were
maintained in a well-ventilated room at a
temperature of 25 ± C with 12/12 h
light/dark cycle in polypropylene cages.
Standard pellet feed and tap water were
provided ad libitum throughout the
experimentation period. The animals were
acclimatized to laboratory conditions for 10
days prior to initiation of experiments. The
project proposal was approved by the
Institutional Animal Ethical Committee (ref.
no. IAEC/A/500/2001).
Acute oral toxicity study
Acute oral toxicity test was performed by
following OECD guideline 420 [10].
Increasing doses of the extracts, up to 2000
mg/kg body wt, were administered to the
mice.
Antitumor studies
Ehrlich ascites carcinoma (EAC) cells were
obtained from the National Centre for Cell
Science, Pune, India and maintained in vivo
in Swiss albino mice, by intraperitoneal (i.p.)
transplantation of 1×10
6
cells/mouse every 10
days. The mice were divided into six groups
of 16 animals each. EAC cells were collected
from the donor mice and suspended in sterile
isotonic saline (0.9 % NaCl). The viable EAC
cells were counted (with the aid of trypan
blue staining) under the microscope and were
adjusted to 10
6
cells/mL. An amount (0.1 mL)
of these cells per 10 g body weight of the
animals was injected (i.p.) into the animals on
day 0. One day of incubation was allowed for
multiplication of the cells.
Ninety six animals were used for study and
they were divided into six groups of 16
animals each. Group I served as control mice
and was administered 0.9 % NaCl solution for
16 days. Group II EAC mice administered
orally chloroform extract (200 mg/kg). Group
III received orally chloroform extract (400
mg/kg); Group IV ethanol extract (200 mg/kg,
orally); Group V ethanol extract (400 mg/kg,
orally); and Group VI the standard drug, 5-
fluorouracil (20 mg/kg, i.p.). The treatments
continued daily for 16 days. On day 21, six
animals from each group were sacrificed and
the remaining animals were kept for further
observation for 3? weeks to determine host
life span.
Determination of antitumor parameters
Tumor volume: The mice were dissected and
the ascitic fluid was collected from the
peritoneal cavity. The fluid volume was
measured in a graduated centrifuge tube and
packed cell volume was determined after
centrifuging (REMI, R4C and REMI Group) at
1000 rpm for 5 min.
Viable/non-viable tumor cell count: The cells
from the preceding test were stained with
trypan blue (0.4 % in normal saline) dye. The
cells that took up the dye were non-viable
while those that did not were viable. Viable
and nonviable cells were counted and cell
count computed as in Eq 1.
Cell count = C/(A x T) …………….. (1)
where C is the number of cells (viable or
nonviable) multiplied by the dilution factor, A
is the area occupied by the liquid film and T is
the thickness of the liquid film.
Increase in life span: The effect of extract on
tumor growth was monitored by recording
mice mortality daily for a period of 6 weeks
from the starting day and the percent
increase in life span (ILS) was calculated for
each animal group with respect to the control
[15-18].
Hematological parameters: At the end of the
experimental period, all the mice were
sacrificed by cervical dislocation. Blood was
collected from a freely flowing tail vein and
used for the estimation hemoglobin (Hb)
content, as well as for red blood cell (RBC)
Chatterjee et al
Trop J Pharm Res, August 2011;10 (4):
450
and white blood cell (WBC) counts. W BC
differential count was carried out using
Leishman stained blood smears [11-17].
Statistical analysis
The results of estimation of biochemical and
functional parameters are reported as mean
value ± standard error of mean (SEM).
Variation within a set of data assessed by
one-way analysis of variance (ANOVA) while
individual comparison of group mean values
by Dunnet’s test (Sigma Stat 3.5). P values <
0.05 were considered statistically significant.
RESULTS
Antitumor activity
The extracts elicited tumor growth response
with respect to packed cell volume, viable
and non-viable cell counts and increase in life
span. Table 1 indicates that the extracts, at
doses of 200 and 400 mg/kg, respectively, as
well as the reference standard drug, 5-
fluorouracil, reduced significantly (p < 0.05)
tumor volume, when compared with the
control group. A similar finding was made for
viable cell count. The number of non-viable
cell count significantly (p < 0.05) increased in
ethanol extract-treated (at a dose of 400
mg/kg) and reference standard groups, but
no significant increase was observed in other
extract-treated groups.
As Table 3 shows, median survival time and
life span of the mice increased following
administration of the extracts at both doses
(200 and 400 mg/kg), thus indicating that the
extracts exhibited a remarkable capacity to
inhibit the growth of solid tumor induced by
EAC cell line in a dose-dependent manner in
experimental animals.
Acute toxicity
The extracts of C. reflexa did not provoke any
gross behavioral changes or manifestations
of toxic symptoms in the animals, such as
weight loss, increased motor activity, tremors,
spasticity, loss of right reflex, decreased
motor activity, ataxia, sedation, muscle
relaxation, hypnosis, arching and rolling,
lacrimation, salivation, watery diarrheoa,
writhing and urination, over an observation
period of 24 h. The extracts were non-lethal
even at the maximum single oral dose of
2000 mg/kg.
Table 1: Antitumor activity of Cuscuta reflexa
extracts with regard to tumor volume, viable tumor
cells count and nonviable tumor cells count
(mean±SEM, n=6)
Treatment
Tumor volume
(ml)
Viable tumor
cells count (10
6
cells/mouse)
Non-viable
tumor cells
count (10
6
cells/mouse)
Control 5.51±
0.38
12.12±
0.45
0.38±
0.10
CHCl
3
extract
(200 mg/kg)
5.11±
0.32
9.60±
0.25*
0.43±
0.05
CHCl
3
extract
(400 mg/kg)
3.98±
0.41*
5.48±
0.41*
0.62±
0.03
Ethanol
extract (200
mg/kg)
4.20±
0.30*
8.13±
0.44*
0.56±
0.04
Ethanol
extract (400
mg/kg)
2.5±
0.0.3
3*
3.06±
0.51*
0.96±
0.11*
5-fluorouracil
(20 mg/kg)
2.31±
0.21*
2.55±
0.34*
0.83±
0.07*
*P <0.05 compared to control group
Hematological effects of the extracts
Table 3 shows that after 16 days of
treatment, the hematological parameters of
the mice were significantly (p < 0.05) altered,
compared to the control group. While total
WBC count increased and Hb content of RBC
decreased in the control group, RBC count
and hemoglobin content increased in extract-
treated groups. Furthermore, there was
decrease in WBC count, as well as counts of
neutrophils and monocytes in extract- and
reference standard-treated groups, while
lymphocyte count rose for these groups.
Chatterjee et al
Trop J Pharm Res, August 2011;10 (4):
451
Table 2: Antitumor activity of Cuscuta reflexa
extracts with regard to median survival time and
increase in life span (mean±SEM, n=10)
Treatment
Median
Survival time
(days)
Increase in
life span
(%)
Control 28.32±2.51 100
CHCl
3
extract
(200 mg/kg) 34.62±1.14 121
CHCl
3
extract
(400 mg/kg) 37.40±2.73 132
Ethanol extract
(200 mg/kg) 37.11±1.58 131
Ethanol extract
(400 mg/kg) 43.26±3.10 152
5-fluorouracil
(20 mg/kg) 46.59±2.43 164
DISCUSSION
The results of the present study indicate the
tumor inhibitory activity of both the chloroform
and ethanol extracts of Cuscuta reflexa
against EAC cells. Usually, the parameters
for evaluating the antitumor activity of a drug
are prolongation of life span, reduction in
tumor volume and improvement in the
hematological parameters of the host.
Ascitic tumor implantation promotes local
inflammatory reactions leading to increase in
vascular permeability, and results in intense
edema formation, cellular migration and
progressive ascitic fluid formation. Ascitic
fluid is essential to tumor growth, since it
constitutes the direct nutritional source for
tumor cells. Decrease in viable cell count and
increase in non-viable cell count in tumor-
bearing mice suggest antitumor activity
against EAC cells in mice [18, 19].
Biological agents, such as interferons and
interleukins, provide non-specific active
immunity (indirect cytotoxic action), whereas
monoclonal antibodies provide passive
immunity (direct cytotoxic action). Interferons
are the small proteins synthesized by the
immune cells in response to various stimuli,
such as viral infections and cytokines, and
inhibit viral replication and promote cellular (T
cell) immune response. Interferons act via
specific cellular receptors linked with JAK-
STAT (Janus Kinase-Signal Transducer
and Activator of Transcription) pathway to
stimulate the formation of specific proteins
which mediate their actions. Their antitumor
action is complex, and includes
antiproliferative effects, promotion of
differentiation, immunomodulation, alteration
Table 3: Effect of Cuscuta reflexa extracts on hematological parameters of Swiss albino mice
(mean ± SEM, n = 6)
Differential count
Treatment Hb content
Total RBC
(cells/ml×10
6
)
Total WBC
(cells/ml×10
6
)
Lymphocyte
(%)
Neutrophil
(%) Monocyte (%)
Control 12.1±0.3 1.29±0.01 17.5±0.28 20.1±3.2 65.2±7.3 4.2±0.5
CHCl
3
extract
(200mg/kg) 12.6±0.2 1.31±0.03 15.17±0.30* 39.5±4.2 53.7±6.2 3.9±0.2
CHCl
3
extract
(400mg/kg) 14.3±0.2* 1.35±0.02 11.2±0.35* 50.6±4.8 32.4±5.8* 3.1±0.4
Ethanol extract
(200mg/kg) 13.5±0.2*
1.36±0.04
12.67±0.27*
43.2±5.1* 42.6±5.4* 3.4±0.5
Ethanol extract
(400mg/kg) 15.9±0.3*
1.45±0.05*
7.03±0.24*
66.6±4.6*
24.5±3.8* 2.8±0.3*
5-fluorouracil
(20 mg/kg) 16.1±0.1* 1.42±0.05* 7.31±0.26* 62.5±3.7* 26.7±7.6* 2.6±0.6*
*P <0.05 compared to control group
Chatterjee et al
Trop J Pharm Res, August 2011;10 (4):
452
in tumor cell surface antigen expression,
inhibition of oncogene activation, and
angiogenesis. The cytokines produced in the
body by the lymphocytes are known as
interleukins and they mediate cytotoxic
actions through the cell surface receptors in
relevant target cells. Interleukins stimulate
the growth and activity of immune cells,
which target cancer cells. It acts as an
antitumor agent by increasing the cytolytic
activity of antigen-specific cytotoxic T
lymphocytes and natural killer (NK) cells and
by increasing the gene expression
responsible for encoding the lytic component
of cytotoxic granules, that is, perforin and
granzymes.
Monoclonal antibodies are the clones of
similar antibodies that are directed against
specific target antigens. Tumor cells express
a wide variety of antigens that are attractive
targets for monoclonal antibody-based
therapy. These antibodies can activate the
immune functions and facilitate the
destruction of malignant cells by
complement-dependent cytotoxicity and
antibody-dependent cell-mediated cytotoxi-
city.
Reduction in viable cell count and increased
non-viable cell count towards normal in tumor
host suggested that extracts stimulate the
growth and activity of immune cells by the
production of Interleukins, which target tumor
cells and cause lysis of the tumor cells by
indirect cytotoxic mechanism. Furthermore,
the reduced volume of EAC and increased
survival time of the mice suggest that the
extracts might have exerted a delay in
vascular permeability to the cells [20]. While
the standard drug, 5-fluorouracil as a
pyrimidine analogue, is transformed inside
the cell into different cytotoxic metabolites
which are then incorporated into DNA and
RNA, eventually inducing cell cycle arrest
and apoptosis by inhibiting the cell's ability to
synthesize DNA. The standard drug activity
thus indicates a gene expression mechanism,
which is indirect cytotoxicity The antitumor
activity of the extracts appear to also follow
this mechanism.
Preliminary phytochemical screening
indicated the presence of flavonoid,
triterpenoids, alkaloids and tannins in the
ethanol extract, and phytosterols,
triterpenoids and alkaloids in the chloroform
extract. It has been reported that flavonoids
possess antimutagenic and antimalignant
effects. Moreover, flavonoids have a
chemopreventive role in cancer through their
effects on signal transduction in cell
proliferation and angiogenesis. The antitumor
properties of the extracts may be due to
these compounds. Phytosterols are able to
be incorporated into the cell membrane,
where they alter membrane fluidity and the
activity of membrane-bound enzymes. They
also alter signal transduction in pathways
leading to tumor growth and stimulate
apoptosis in tumor cell lines [21,22]. This
supports the indirect cytotoxic mechanism of
both extracts.
The results also revealed that the ethanol
extract showed significantly higher antitumor
activity than the chloroform extract; this may
be due to the presence of triterpenoids,
alkaloids and tannins which might have
enhanced the antitumor property of the
flavonoids in the extract. On the other hand,
triterpenoids and alkaloids in the chloroform
extract appear not to have enhanced the
antitumor property of phytosterols in the
extract.
Carcinogenesis or tumor development
generates highly toxic and diffusible reactive
oxygen species that cause extensive cell
damage or adducts on the biomolecules,
contributing to malignant transformation.
Erythrocytes membranes are particularly
susceptible to oxidative stress due to its high
content of polyunsaturated fatty acids, which
are more vulnerable to oxidative damages.
The erythrocytes are particular susceptible to
peroxide stress due to high content of iron, a
potent catalyst for the production of reactive
oxygen species and due to continuous
Chatterjee et al
Trop J Pharm Res, August 2011;10 (4):
453
challenge with high oxygen tension [23].
Susceptibility of erythrocytes to peroxides
was confirmed by the elevated WBC count,
reduced hemoglobin and RBC count, as
observed in the control group. The
constituents of the extracts probably
facilitated improvement in hematological
parameters by scavenging reactive oxygen
species and improving the antioxidant
defense system, thus indicating that the
extracts exerted haematopoietic protective
activity as well as antioxidant property
[24,25].
CONCLUSION
The chloroform and ethanol extract of
Cuscuta reflexa possess significant antitumor
activity and increased the life span of tumor-
bearing mice. Further investigations are in
progress in our laboratory to identity the
active principles involved in this antitumor
activity.
ACKNOWLEDGEMENT
The authors are thankful to Dr RC Agrawal,
Asst. Professor, Jawaharlal Nehru Cancer
Hospital & Research Centre, Bhopal (M.P.)
for making available laboratory facilities for
the work and giving guidance in the
pharmacological aspects of the studies.
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... The inhibition of spontaneous mobility of earthworm was shown by extract off C. reflexa. Anthelmintic activity was also shown by chloroform and methanolic extract when compared with albendazole (89) Anticancer Activity/ Antiproliferative Activity Significant anti-tumor activity was seen by chloroform and ethanol extract in Ehrlich ascites carcinoma bearing mice, with standard 5fluorouracil (90). ...
... The chloroform and ethanol extracts of C. reflexa in EAC-bearing mice have shown a positive antiproliferative effect (90). ...
... The drug has several pharmacological active constituents, and several studies have been done to validate its uses in various literature, including USM. Its antioxidant (100), anticancer (91,92,90), antiproliferative (81,93), anticonvulsant (94), anti-HIV activities (43) have been proved by various pharmacological studies and showed promising results. The results support its importance and usefulness in the treatment of diseases like cancer. ...
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Cuscuta reflexa Roxb. (Fam. Convolvulaceae) is commonly known as aftimoon, amarbel, akashbel, or dodder in the alternative medicine system. It is widely used in the Unani system of medicine (USM) for its useful therapeutic effects due to its active constituents. It is used to cure various diseases through anti-tumor activity, anti-arthritis, hypotensive and bradycardia effects. Various phytoconstituents were isolated from it, like protein, alkaloids, flavonoids, dulcitol, luteolin, quercetin. The kaempferol and quercetin, a major active constituent in it, have various therapeutic activities apart from antidiabetic activity. In USM, Cuscuta reflexa is used to treat hepatitis, melancholia, chronic splenitis, and cancer. Many pharmacological studies have been carried out to explore its hypotensive, anti-HIV, antibacterial, antioxidant activities. This review will provide a comprehensive overview of Cuscuta reflexa with special reference to Unani medicine and validate its various effects by pharmacological studies done so far.
... It has been used historically as a purgative, demulcent, diaphoretic, and therapy for prolonged fever [3,4]. According to numerous pharmacological studies, this plant is used to treat a number of ailments, including alopecia [5], HIV [6], 1997), diabetes [7], epilepsy [8], CNS depression, pain [9], tumour [10], and urinary infections [11]. It also acts as an antihistaminic, anticholinergic, anti-hypertensive [12], antibacterial, antioxidant, anthelmintic, and antiinflammatory agent [13][14][15]. ...
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The antimicrobial effect of Cuscuta reflexa Roxb. (Cuscutaceae) stem extract was evaluated on microbial strains like Gram positive species Staphylococcus aureus and Bacillus subtilis and Gram negative species Escherichia coli and Pseudomonas aeruginosa. Petroleum ether, chloroform, and alcohol were the solvents used for plant extraction. The stem's alcoholic extract exhibits the highest level of antibacterial activity. The disc diffusion method was used to assess in vitro antibacterial activity. Amphicillin, a common antibiotic, and the substantial antibacterial activity of the active extract were compared. The phytochemical studies also utilized the stem sample material. The results of the phytochemical analysis showed the presence of alkaloids, oil, fat glycosides, carbohydrates, phenolics, tannins, lignin, saponins, flavonoids, and terpinoids in the sample. The stem's antibacterial activities are due to the presence of several secondary metabolites. The standard methods was used to assess the phytochemical screening and antibacterial activity.
... C. reflexa is known as a wonder plant, which explains why a large number of pharmacological studies have been conducted on it, utilizing various exploration techniques and investigations (Durdana et al., 2010). These pharmacological examinations show the utilization of this plant as an antioxidant and antifungal (Parveen et al., 2013), anti-inflammatory, lower blood pressure, antibacterial (Mala and Sofi, 2017), for the treatment of antihistamine, alopecia (Ambi et al., 2017), anticonvulsant, anti-HIV, anticholinergic, antidiabetic (Mahmood et al., 1997Rahmatullah et al., 2010), muscle relaxant, analgesic, antitumor, diuretic, CNS depression, and anthelmintic (Chatterjee et al., 2011;Udavant et al., 2012), It also has anti-inflammatory and anti-cancer activity (Suresh et al., 2011). Various distributed articles have detailed the antidiarrheal employment of C. reflexa by various networks, albeit test proof is missing (Mukherjee et al., 2006;Khan et al., 2016;Sharma and Kapoor, 2014). ...
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Cuscuta reflexa is an extensive leafless, parasitic climber that has been used since ancient times for various purposes and as a therapeutic plant in different areas of Pakistan. The phytochemical, antibacterial, antioxidant, and fatty acid investigations were carried out on the leaves, stems, and fruit extracts of C. reflexa. Preliminary phytochemical screening has shown the presence of various phytochemicals such as carbohydrates, protein, amino acids, alkaloids, flavonoids, phenols, glycosides, saponins, tannins, terpenoids, steroids, and phenolic compounds. The percentage variation of some components fluctuated like moisture (6.85-10.34%), ash (5.38-7.83%), acid insoluble ash (0.28-0.71%), water-soluble ash (0.78-0.96%), hexane extractives (1.23-1.91%), alcohol extractives (10.39-12.23%), water extractives (25.34-30.35%), loss on drying (1.16-1.47%) and crude fiber (15.04-18.26%) for leaves, stems, and fruits respectively. The level of free fatty acid was different as follows: leaves (2.37%), stems (2.16%), and fruits (2.94%); a high value of peroxide was observed in stems, followed by leaves and fruits. The iodine and saponification values in leaves, stems, and fruits were found in the range of 105-116 g I2/100 g, and 165-175 mg KOH/g. The antibacterial activity was carried out by the disc diffusion method against gram-positive Staphylococcus aureus and gram-negative Escherichia coli bacteria. It was found that all extracts of the selected plant were most active against E. coli with a zone of inhibition of 4, 4, and 6 at MIC 250 as compared to S. aureus, which showed a zone of inhibition of 3, 4, and 5 at MIC 250 µg/ml. The antioxidant potential of the leaves, stems, and fruit extracts, examined on the basis of their scavenging activity of free radicals, was found to be good. The result of the gas chromatography-mass spectroscopy (GC-MS) analysis reveals that linoleic acid (38.10-40.53%) is the most abundant fatty acid of C. reflexa and the smallest amount was found in tricosanoic acid (0.12-0.19%). In general, the isolated compounds were reported to possess anticarcinogenic, antitumor, antimicrobial, and anti-inflammatory properties. It was concluded that C. reflexa has a markedly therapeutic potential to heal wounds and may provide the pharmacological basis for AcademicPres Notulae Botanicae Horti Cluj-Napoca Agrobotanici Sidhu AR et al. (2022). Not Bot Horti Agrobo 50(3):12691 2 its folk uses.
... 76 Cuscuta reflexa (Cuscutaceae): Chloroform and ethanol extracts acted against Ehrlich Ascites Carcinoma (EAC) cell line. 77 Cuscutareflexa has shown anticancer activity against leukemias and melanoma. 78 It has also shown anticancer activity on Hep3B cells by up-regulation of pro-apoptotic factors BAX and p53and downregulation of anti-apoptotic factors Bcl-2 and surviving. ...
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According to WHO, cancer is one of the leading causes of death worldwide, accounting for an estimated 9.6 million deaths in 2018. More significant improvements have been made in the management and treatment of cancer, still there remains scope for the betterment of treatment procedures. Mostsynthetic anticancer drugs are known to develop resistance, show cytotoxicity against normal cells due to their non-selective nature, and cause tremendous side effects. Medicinal plants are significantly feasible sources of organic compounds, for their better availability, cheaper price, fewer side effects, and sometimes better therapeutic efficacy, which may benefit the world commercially or act as an important starting point for identifying lead compounds to develop modified derivatives. This article describes the ethnobotanical properties of 15 available medicinal plants of Bangladesh having anti-cancer properties.
... It induced apoptosis in Hep3B cells through the up-regulation of p53, BAX, and downregulation of Bcl-2 and surviving [19] . One another study documented that, chloroform and ethanol extracts of C. reflexa exhibit significant antitumor activity in EAC-bearing mice that is comparable to that of the reference standard, 5-fluorouracil [20] . ...
... However, tris (dimethylamino)phosphine (13) is a strong nucleophile [51] used to synthesize epoxides and arene oxides from aldehydes [52][53][54]. Moreover, it replaces triphenylphosphane in the Wittig reaction for the preparation of unsaturated esters as well as alkenes [55] and provides convenient synthetic access to epoxides [55,56]. So the reaction of tris (dimethylamino)phosphine (13) with 4-methoxyphenylisothiocyanate (1) was conducted in THF. ...
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Phosphonium compounds offer an attractive branch of research that chemists and biologists apply for producing many novel drugs for various applications, and its polymeric ingredients are composed of quaternary ammonium and phosphonium salts. The reactions of isothiocyanate with phosphinimine bestow thiaziridine, carbamate, and thiourea derivatives. Moreover, isothiocyanate reacts with tris (dimethylamino) phosphine leading to the formation of sulfidomethyl phosphonium. Lawesson's and Japanese reagents have potential to react with isothiocyanates to generate dithiaphosphetane sulfides. Treatment of isocyanate with Lawesson's or Japanese reagents under reflux conditions affords thiaphosphetidinone sulfide, but when applied at room temperature the dithiaphosphetane sulfide was isolated. Ehrlich ascites carcinoma (EAC) mice model was used to investigate potential anticancer properties of the novel phosphonium and thiophosphate derivatives. Synthesized compounds (100mg/kg b.w.) were administered orally to the EAC-bearing mice for about two weeks. Compounds’ antineoplastic activity was determined by the evaluation of volume, viability, and inhibition percent of EAC cells. In addition, DNA fragmentation percent was assessed. The expression of apoptotic marker genes (Bax, Bcl2, Caspase 3) and encoding proinflammatory cytokines (TNF-α) and pro-apoptotic protein (p53) were inspected by RT-qPCR. The overall conclusion was based on the findings that treatment with synthesized compounds leads to decrease in tumor volume; increase in tissue DNA fragmentation, downregulation of Bcl2 gene, and up-regulation of Bax, caspase3, and p53 markers, along with decrease in TNF-α level in liver tissues. These findings suggest that the anticancer mechanism of these compounds is based on the programmed cell death (Apoptosis).
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Introduction This study is an effort to document extensively and systematically the ethnobotanical and ethnomedicinal knowledge in the four districts (Puri, Cuttack, Bhadrak, and Mayurbhanj) of Odisha in Eastern India. It provides new insights into the rich ethnomedicinal knowledge and plants biodiversity in these four districts. Methods The method of convenience sampling was conducted to get the ethnobotanical and ethnomedicinal knowledge of the healers. Semi-structured interviews were conducted, and the information got was quantified and documented with the aid of various databases. Eleven traditional healers endowed information on their traditional medicinal knowledge. 74 plant species belonging to 44 botanical families are documented and among these 44 botanical families, five (Rutaceae, Malvaceae, Astaraceae, Annonaceae, and Euphorbiaceae) are mostly used by the healers. The healers mainly use leaves and roots in the medicine's preparation. Results Twenty-five ailments are treated by these traditional healers, which are majorly dominated by dermatological treatments. The pharmacological mining of these 74 plants revealed that a few pharmacological and biological activities of each medicinal plant have been studied. Our study revealed that these four districts of Odisha have a rich biodiversity of medicinal plants. Conclusion Promoting the plantation of some of these species can enhance the income of the traditional healers of these districts and simultaneously make the exploitation of these plant species sustainable. We also found that the pharmacological and biological activities of many plant species are yet to be explored.
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The purpose of this study is to determine whether the complexing hydroalcoholic extract of Cuscuta reflexa (HECR) with phosphatidyl choline increases its bioavailability. As a result, a novel phytosomal delivery system for the HECR-soya lecithin complex was developed (HECR-phytosome). The HECR-phytosome complex was synthesized and characterized as phytovesicles. The formulation was prepared using a variable concentration of soya lecithin (1:1-1:3 percent w/v), a temperature range of (45-65°C), and sonication time (4-8 min). Optimization of HECR-loaded phytosomal formulations was performed using Design Expert software. A three-factor, three-level Box-Behnken design was used to optimize this HECR delivery system, as dependent variables, vesicular size and entrapment efficiency were evaluated using a Box Behnken factorial design. Further characterization of the optimized formulation included vesicle size, PDI, zeta potential, entrapment efficiency, FTIR, DSC, TEM, and in vitro release. Vesicle sizes ranged from 173.5±6.17 nm to 215.9±6.53 nm, and response rates for entrapment efficiency ranged from 52.9±1.65 to 77.2±1.1%. The uniform structure and spherical shape were demonstrated by transmission electron microscopy. Among the drug release kinetic models, the formulation followed the Higuchi model (R² = 0.9978), releasing 96.3±3.7% of the polyphenol and flavonoids phytoconstituents from HECR-loaded phytosomes in 12 hours, compared to 49.3±2.5% in the plain extract. In addition, the optimized formulation passes the stability test. Therefore, the results demonstrated that phytosomal nanocarriers have the potential to increase the bioavailability of Cuscuta reflexa extract. graphical abstract Fullsize Image
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Introduction The consumption of ‘Patalagarudi’ (Cocculus hirsutus ‘CHP’) and ‘Amarbel’ (Cuscuta reflexa ‘CRA’) as ethnic plants for health promotions rarely validated. The limited literature reported these plants as antioxidant and immunomodulators. Objective To evaluate the biodynamic properties of CHP and CRA extracts. Methodology The traditional formulation, ‘Kwath’ (K) and conventional extracts were prepared with CRA and CHP. The total phenolic content (TPC) was estimated. Various polyphenol compounds in the extracts were eluted on UHPLC. The biodynamic activities; i. Free radical scavenging (FRS-DPPH and ABTS), ii. Intracellular ROS scavenging activity in RAW 264.7 cell line iii. Spleenocytes proliferation assay for Th1/Th2 Immunomodulatory potential by flow-cytometer were assessed. Results The TPC in CRA (105–159 μg GAE/mg) and CHP (35–48 μg GAE/mg) recorded. The chromatographic peaks confirmed the presence of polyphenols in CRA and CHP extracts. UV spectra of the extracts to the extent possible have been correlated with certain polyphenols. The FRS (IC50) was significantly low in CRA-K (DPPH = 22.7; ABTS = 12.0 μg/ml) than CHP-K (DPPH = 70.4; ABTS = 50.2 μg/ml). Similarly, intracellular ROS scavenging activity with CRA-K (84%) showed the highest inhibitory potential compared to CHP-K (50%) and LPS control. The immunomodulatory activity of CRA-K significantly upregulated TH1 cytokines (TNFα and IFN-γ). The downregulation of Th2 cytokines (IL-4 and IL-10) was in all CRA and CHP extracts as compared to Con A. Conclusion The current study confirms the immunomodulatory and antioxidant properties of CRA and CHP along with the presence of polyphenols.
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Ethnopharmacological relevance Cuscuta reflexa Roxb. (C. reflexa) is a well-known traditional herbal plant, with numerous inherent therapeutic potentials including anticancer, antitumor, antibacterial, analgesic, anthelmintic, laxative and others. Moreover, the anticancer and antitumor potentials of this herb are ongoing with several trails, thus an attempt was made to assess the anticancer and hepatoprotective potentials of traditional C. reflexa herbs. Method The dried ethanolic extract of C. reflexa was tested for acute oral toxicity in the treated animals subsequently their behavioral, neurological, and autonomic profiles changes were observed. The preliminary anti-cancer effects of extracts against 1, 2- Dimethyl hydrazine (DMH) induced animals were assessed through barium enema X-ray, colonoscopy, and Aberrant crypt foci (ACF) studies. The blood samples of the animals (treated and untreated) were collected and their in-vitro histological parameters were evaluated by the experienced technician. Results It was observed that C. reflexa significantly reduced Disease activity indexing (DAI) level and ACF counting, as well as demonstrated similar activity as of the standard drug 5-Fluorouracil (5-FU). Histopathological results revealed that the apoptotic bodies decreased in the DMH-induced group (group II) during cancer progression while in 5-FU treated (group III) and C. reflexa treated (group IV and V) animals the apoptotic bodies were increased. Inversely, the mitotic bodies increased in group II animals and reduced in group III, IV, and V animals. In the colonic section, DMH-induced cancer assay exhibited significant effects on the levels of hemoglobin, Packed cell volume (PCV), Red blood cell (RBC) counts, Mean corpuscular hemoglobin concentration (MCHC), Mean corpuscular volume (MCV), and Mean cell hemoglobin (MCH), and was found to be less in group II animals whereas administration of C. reflexa efficiently recovered back the loss probably by healing the colon damage/depletion of cancer progression. Moreover, compared to the group II animals, the neutrophil count was within the normal range in C. reflexa administered group. Conclusions In the present study, the major hematological parameters significantly increased within DMH treated animals and exhibited extensive damage in the hepatic regions. Moreover, the histopathological findings demonstrated that the C. reflexa extracts potentially reduced the cell proliferation, with no toxicity. The C. reflexa extracts exhibited impending anti-cancer activity as well as protected the hepatic cells and thus could be potentially used in the management of colon or colorectal cancer and hepatic impairments.
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The present study was carried out to evaluate the antitumor potentials of Amaranthus spinosus against EAC bearing Swiss albino mice. The ethanol extract of its leaves given orally to mice at the dose of 100 and 200 mg/kg body weight for 16 days. It was observed that decrease in tumor volume and viable cell count, while increase in mean survival time and non viable tumor cell count, when compared to the mice of the EAC control group. Restoration of hematological and biochemical parameters towards normal was also observed. The results suggest that the ethanol extract of Amaranthus spinosus leaves exhibits significant antitumor effects in EAC bearing mice.
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Two new phenylpropanoid diglucosides named syringinoside and coniferinoside have been isolated from the water-soluble fraction of Wikstroemia sikokiana FRANCE. et SAV. (Thymelaeaceae). Their structures have been characterized on the basis of spectral and chemical evidence.
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The objective of the present study is to explore the anticancer activity of the ethanolic and aqueous extracts of the Dendrophthoe falcata in Swiss albino mice against Ehrlich Ascites Carcinoma (EAC) cell line. Anticancer activity of ethanolic and aqueous extracts of D. falcata was evaluated in EAC Swiss albino mice at the doses of 200 and 400 mg/kg body weight orally. Both extracts at both doses were administered for 13 consecutive days. After 24 h of the last dose and then eighteen hours of fasting, the mice were sacrificed and antitumor effect of ethanolic and aqueous extracts was as- sessed by evaluating tumor volume, viable and nonviable tumor cell count, tumor weight and hematological parameters of EAC bearing host. Ethanolic and aqueous extracts showed significant decrease in (p<0.0001) tumor volume, viable cell count, tumor weight and elevated the life span of EAC tumor bearing mice. Haematological profiles such as red blood cell (RBC), haemoglobin, and white blood cell (WBC) count reverted to normal level in treated mice. The results demon- strated that the extract has potent dose dependent anticancer activity comparable to that of cisplatin. Aqueous extract at both doses (200 and 400 mg/kg) and ethanolic extract at 400 mg/kg dose showed potent anticancer activity.